Total Synthesis of Psoralidin, an Anticancer Natural Product - The

Mar 2, 2009 - A base-catalyzed condensation of phenyl acetate with acid chloride, followed by intramolecular cyclization and microwave-assisted cross-...
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Total Synthesis of Psoralidin, an Anticancer Natural Product Pallab Pahari and Ju¨rgen Rohr* Department of Pharmaceutical Sciences, College of Pharmacy, UniVersity of Kentucky, 725 Rose Street, Lexington, Kentucky 40536-0082 [email protected] ReceiVed NoVember 21, 2008

A base-catalyzed condensation of phenyl acetate with acid chloride, followed by intramolecular cyclization and microwave-assisted cross-metathesis reaction, leads to the first total synthesis of psoralidin, a natural product with a broad range of biological activities, in a highly convergent and regioselective manner.

Introduction Psoralidin (1) was first isolated from seeds of Psoralea corylifolia Linn in 1948 by Chakravarti et al.,1 and later its structure was corrected by Khastgir et al. in 1961.2 It was found to be an active ingredient of many Indian and Chinese traditional herbal medicines.3 Psoralidin (1) exhibits a variety of biological activities like antioxidant,4 antibacterial,5 and antidepressant activities.6,7 It also shows inhibitory activities against protein tyrosine phosphatase 1B, which plays a major role in the negative regulation of insulin signaling.8 In addition, psoralidin inhibits antigen IgE-induced degranulation in RBL-2H3 cells9 and acts as a potent antidepressant by strongly inhibiting forskolin-induced corticotrophin releasing factor (CRF) gene transcription.6,7 Psoralidin also has great potential as an anticancer agent, showing cytotoxic effects on gastric (SNU-1 and SNU-16), colon (HT-29), and breast (MCF-7) cancer (1) Chakravarti, K. K.; Bose, A. K.; Siddiqui, S. J. Sci. Ind. Res. 1948, 7B, 24–26. (2) Khastgir, H. N.; Sengupta, P.; Duttagupta, P. C. Tetrahedron 1961, 14, 275–283. (3) Ranga, R. S.; Girija, R.; Nur-e-alam, M.; Sathishkumar, S.; Akbarsha, M. A.; Thirugnanam, S.; Rohr, J.; Ahmed, M. M.; Chendil, D. Cancer Chemother. Pharmacol. 2004, 54, 7–15. (4) Guo, J. M.; Weng, X. C.; Wu, H.; Li, Q. H.; Bi, K. S. Food Chem. 2005, 91, 287–292. (5) Khatune, N. A.; Islam, M. E.; Haque, M. E.; Khondkar, P.; Rahman, M. M. Fitoterapia 2004, 75, 228–230. (6) Yi, L. T.; Li, Y. C.; Pan, Y.; Li, H. M.; Xu, Q.; Mo, S. F.; Qiao, C. F.; Jiang, F. X.; Xu, H. X.; Lu, X. B.; Kong, L. D.; Kung, H. F. Prog. NeuroPsychopharmacol. Biol. Psychiatry 2008, 32, 510–519. (7) Chen, Y. C.; Cheung, Y. T.; Kong, L. D.; Ng, T. B.; Qiao, C. F.; Mo, S. F.; Xu, H. X.; Kung, H. F. Life Sci. 2008, 82, 1117–1121. (8) Kim, Y. C.; Oh, H.; Kim, B. S.; Kang, T. H.; Ko, E. K.; Han, Y. M.; Kim, B. Y.; Ahn, J. S. Planta Med. 2005, 71, 87–89. (9) Matsuda, H.; Sugimoto, S.; Morikawa, T.; Matsuhira, K.; Mizuguchi, E.; Nakamura, S.; Yoshikawa, M. Chem. Pharm. Bull. 2007, 55, 106–110.

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FIGURE 1. Chemical structures of psoralidin and related natural products.

cell lines.10,11 Recently, we reported that 1 inhibits protein kinase Akt phosphorylation, and thus, inhibits the growth of androgenindependent prostate cancer cells both in Vitro and in ViVo.12 Psoralidin 1 is a member of the coumestane family of natural products having an isopentenyl group at the C-2 position of coumesterol (4). In 1961, Nashipuri and Pyne13 reported the synthesis of dihydropsoralidin (2) and confirmed the structure of the natural product. Jain et al.14 reported the synthesis of some prenylated coumestanes as analogues of psoralidin, and most recently Fu¨rstner et al.15 synthesized and corrected the proposed structure of phaseol (3), a structural isomer of 1. (10) Mar, W.; Je, K. H.; Seo, E. K. Arch. Pharmacal Res. 2001, 24, 211– 213. (11) Yang, Y. M.; Hyun, J. W.; Sung, M. S.; Chung, H. S.; Kim, B. K.; Paik, W. H.; Kang, S. S.; Park, J. G. Planta Med. 1996, 62, 353–354. (12) Kumar, R.; Srinivasan, S.; Koduru, S.; Pahari, P.; Rohr, J.; Kyprianou, N.; Damodaran, C. Cancer PreV. Res. 2009, 2, in press. (13) Nasipuri, D.; Pyne, G. J. Chem. Soc. 1962, 3105–3106. (14) Jain, A. C.; Jain, S. M.; Gupta, S. M.; Singh, K. V. Indian J. Chem., Sect. B 1984, 23, 1028–1029. (15) Fu¨rstner, A.; Heilmann, E. K.; Davies, P. W. Angew. Chem., Int. Ed. 2007, 46, 4760–4763.

10.1021/jo8025884 CCC: $40.75  2009 American Chemical Society Published on Web 03/02/2009

Total Synthesis of Psoralidin SCHEME 1.

Retrosynthetic Analysis

However, there are no reports on the synthesis of 1 so far. In our ongoing work on the investigation of the chemical constituents of Rasagenthi lehyam, a complementary and alternative medicine for prostate cancer used as Ayurveda and Siddha medicine in different parts of India, we isolated 1 as the active principle of this mixture of 38 different botanicals, metals, and fats.12 However, the isolated natural product was found to be insoluble in water and, therefore, less effective for in ViVo studies. To overcome this problem, we plan to generate a synthetic library of psoralidin analogues that can be utilized to study structure-activity relationships (SAR), which in turn will pave the way for the synthesis of an active analogue or derivative with increased water solubility. Herein, we report an efficient and general synthesis of 1 that can be easily adopted for the synthesis of various analogues. Many routes to prepare coumestanes can be found in literature, but most of them are complicated and require multistep syntheses.16-24 Retrosynthetically, 1 can be reduced to 5 through the opening of the lactone and furan rings; in turn, 5 can be obtained from two relatively simple starting materials, namely 2,4-dimethoxy-phenylacetic acid methylester (6) and 2,4-dimethoxy-5-prenyl benzoylchloride (7) (Scheme 1). This strategy was expected to be flexible enough for the preparation of various analogues, considering the facile preparation of the starting materials. Al-Maharik and Botting24 used a similar strategy for their synthesis of 13C-labeled 4.

SCHEME 2

SCHEME 3

Results and Discussion We started our synthesis of 1,5-dibromo-2,4-dimethoxybenzene (9),25 which was prepared from resorcinol 8 through bromination with Br2/CHCl3 and O-methylation of the hydroxyl groups using K2CO3/MeI. Br/Mg exchange with i-PrMgCl at ∼10 °C and quenching with prenylbromide provided 10 exclusively in 65% yield. The same reaction was carried out (16) Chang, C. F.; Yang, L. Y.; Chang, S. W.; Fang, Y. T.; Lee, Y. J. Tetrahedron 2008, 64, 3661–3666. (17) Li, C. C.; Xie, Z. X.; Zhang, Y. D.; Chen, J. H.; Yang, Z. J. Org. Chem. 2003, 68, 8500–8504. (18) Takeda, N.; Miyata, O.; Naito, T. Eur. J. Org. Chem. 2007, 149, 1501– 1509. (19) Yao, T. L.; Yue, D. W.; Larock, R. C. J. Org. Chem. 2005, 70, 9985– 9989. (20) Hiroya, K.; Suzuki, N.; Yasuhara, A.; Egawa, Y.; Kasano, A.; Sakamoto, T. J. Chem. Soc., Perkin Trans. 1 2000, 4339–4346. (21) Kraus, G. A.; Zhang, N. J. Org. Chem. 2000, 65, 5644–5646. (22) Mali, R. S.; Tilve, S. G. Synth. Commun. 1990, 20, 1781–1791. (23) Pandey, G.; Muralikrishna, C.; Bhalerao, U. T. Tetrahedron 1989, 45, 6867–6874. (24) Al-Maharik, N.; Botting, N. P. Tetrahedron 2004, 60, 1637–1642. (25) Srebnik, M.; Mechoulam, R.; Yona, I. J. Chem. Soc., Perkin Trans. 1 1987, 1423–1427.

using n-BuLi and diprenylated and monodebrominated side products were found that were not easy to separate. Acid 11 was obtained by replacing the bromine atom of 10 using n-BuLi and CO2 gas that was bubbled through the reaction mixture. The other starting molecule, compound 6, was prepared by the condensation of 2,4-dimethoxybenzaldehyde and hippuric acid following a literature procedure.26 Treatment of acid 11 with oxalyl chloride in the presence of catalytic amounts of DMF produced acid chloride 7 that, without further purification, was reacted with the anion obtained from the treatment of 6 with J. Org. Chem. Vol. 74, No. 7, 2009 2751

Pahari and Rohr SCHEME 4

LDA and produced the desired compound 5 in an overall 82% yield. The structure of 5 was confirmed by spectroscopic analysis. At this point, it was expected that demethylation followed by intramolecular cyclization would yield 1 in a one pot reaction. The demethylation of the four methoxy groups was carried out with BBr3 at 0 °C, but the attempted in situ cyclization ended up with isopsoralidin (12) instead of 1.2 The structure of 12 was confirmed by comparison with reported data (Scheme 2).13,27 The formation of 12 can be explained by the generation of a stable tertiary carbocation produced from the prenyl group in acidic conditions, which is attacked by the orthohydroxy group to yield the undesired dihydropyran ring. A similar reaction of a prenyl group was reported by Molyneux and Jurd.27 To avoid this undesired reaction, we modified our strategy and planned to introduce the prenyl group at the concluding step of the reaction sequence. Thus, we started with acid 13,28 which was prepared from 9 by treatment with n-BuLi/CO2 (Scheme 3). The reaction was optimized using less than one equiv of n-BuLi and provided a single product 13 in 81% yield. Reaction of the corresponding acid chloride 14 with 6 under similar conditions as above provided 15, which was demethylated and cyclized into 16 in 78% yield. The structures of compounds 13-16 were confirmed by 1H and 13C NMR spectroscopy, as well as HRMS. Protection of the two phenolic OH-groups of 16 as MOM-ethers, using diisopropylethylamine (i-Pr2NEt) and methoxy methyl chloride (MOMCl), gave compound 17 in 65% yield. It was anticipated that Br/Mg or Br/Li exchange of 17 followed by electrophilic addition of prenyl bromide would complete the synthesis. However, all attempts to convert 17 with i-PrMgBr/prenyl bromide or n-BuLi/ prenyl bromide under various reaction conditions ended up with complex product mixtures. A Pd(0)-catalyzed Ullmann type cross-coupling reaction also failed to give any product. (26) Barua, A. K.; Chakrabarty, M.; Datta, P. K.; Ray, S. Phytochemistry 1988, 27, 3259–3261. (27) Molyneux, J. R.; Jurd, L. Tetrahedron 1970, 26, 4743–4751. (28) Rick, G. P. J. Am. Chem. Soc. 1926, 48, 3125–3130.

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In the search for a specific functional group which could survive the acidic conditions necessary for the cyclization reaction and could be easily transformed to a prenyl group, we considered the olefin cross-metathesis reaction. Recently Hastings et al.29 used an olefin cross-metathesis reaction in the presence of Grubbs second-generation catalyst to convert an allyl side chain to prenylated compound in very good yields. We hoped that a possible primary or secondary carbocation produced from an allyl group under acidic conditions would not be stable enough to cause an undesired cyclization reaction, like in the case of the tertiary carbocation from the prenyl residue described above. We prepared the allylated compound 18 from 9 using i-PrMgBr and allyl bromide. A similar reaction sequence using n-BuLi/CO2 and 6/LDA as described above provided compounds 19 and 21, respectively. Demethylation and cyclization produced the expected allylated compound 22 in 85% yield along with very small amounts of the dibrominated side product 23 (1%, Scheme 4). Compounds 18, 19,and 21-23 were characterized by 1H and 13C NMR spectroscopy, as well as HRMS for structure verification. Our attempt to perform the alkene cross-metathesis reaction of 22 in the presence of a Grubbs second-generation catalyst and 2-methyl-2-butene initially failed when using identical reaction conditions as described by Hastings et al.29 Even stirring the reaction at rt for 72 h or refluxing the mixture for 48 h with repeated addition of Grubbs catalyst and 2-methyl-2-butene failed to give any isolable product. To check the feasibility of the reaction, we performed the same reaction on 18 which produced prenylated compound 10 in very good yields. Since compound 22 might require a higher temperature for the crossmetathesis reaction, which was limited in this case by both the chosen solvent and the reagent 2-methyl-2-butene, we decided to carry out the reaction in a sealed tube in a microwave oven to allow higher reaction temperature and to avoid loss of starting material.30 Thus, the final step of the total synthesis was (29) Hastings, J. M.; Hadden, M. K.; Blagg, B. S. J. J. Org. Chem. 2008, 73, 369–373.

Total Synthesis of Psoralidin

achieved by heating a mixture of 22, Grubbs second-generation catalyst, and 2-methyl-2-butene in CH2Cl2 in a sealed tube microwave reactor at 100 °C for 15 min. The reaction produced 1 in 73% yield along with compound 24 as a minor side product. All spectroscopic data of the synthetic compound matched those of the natural product (Table 1 in Supporting Information) except for the melting point, which was significantly higher for the synthetic compound than reported for the natural product. Since there was no ambiguity about the structure of the synthetic compound, we assumed that the melting point of the natural product was lowered by minor impurities. Conclusions In summary, we achieved the first total synthesis of psoralidin 1 in a convergent and highly regioselective manner. The synthetic scheme described here is straightforward and efficient. The longest linear sequence is only five steps starting with commercially available resorcinol. The process is readily adaptable and flexible as substitutions can be introduced in both the phenyl acetate and the allyl benzoyl chloride starter molecules to furnish numerous analogues for SAR studies. Further structural diversification could be achieved on the level of the concluding Grubbs reaction. Biological assays to determine and compare the efficacy of 16, 22, and 23 with that of 1, as well as syntheses of various regioisomers, are in progress. Experimental Section31 Psoralidin 1. Grubbs second-generation catalyst (3 mg, 0.0035 mmol) was added to a degassed solution of 22 (20 mg, 0.06 mmol) in CH2Cl2 (10 mL) and 2-methyl-2-butene (2 mL). The solution was heated in a sealed tube at 100 °C for 15 min by a microwave reactor (Biotage Initiator 2.0, standard configuration, temperature control, external IR temperature sensor, fixed hold time). CH2Cl2 was removed in vacuum, and the mixture was purified by reverse phase semipreparative HPLC (symmetry 18C, 7 µm, solvent gradient 9:1 CH3CN:H2O) to get compound 1 (16 mg, 73%) as a white solid along with a minor amount of side product 24 (1 mg, 5%). Rf ) 0.4 (50% ethyl acetate in hexane); mp >400 °C (lit.1 315 °C); νmax (film) cm-1 1719, 1629, 1597, 1419, 1370, 1260, 1092. 1H NMR (DMSO-d6, 500 MHz) δ 7.68 (d, 1H, J ) 8.0 Hz), 7.62 (s, 1H), 7.17 (d, 1H, J ) 2.0 Hz), 6.94 (dd, 1H, J ) 8.0, 2.0 Hz), 6.93 (s, 1H), 5.35 (t, 1H, J ) 7.5 Hz), 3.32 (d, 2H, J ) 7.5 Hz), 1.74 (s, 3H), 1.70 (s, 3H). 13C NMR (DMSO-d6, 125 MHz) δ 159.6 (C), 159.0 (C), 157.8 (C), 157.0 (C), 156.0 (C), 152.9 (C), 132.6 (C), 126.5 (C), 121.8 (CH), 121.0 (CH), 120.6 (CH), 114.7 (C), 114.0 (CH), 103.8 (C), 102.4 (CH), 102.0 (C), 98.8 (CH), 27.6 (CH2), 25.7 (CH3), 17.7 (CH3). HRMS (EI+) m/z 336.1008 ([M]+ C20H16O5, requires 336.0998). (E)-2-(But-2-enyl)-3,9-dihydroxy-6H-benzofuro[3,2-c]chromen-6-one (24). Rf ) 0.35 (50% ethyl acetate in hexane); mp >400 °C; νmax (film) cm-1 1718, 1637, 1629, 1267, 1213, 1093. 1H NMR (DMSO-d6, 500 MHz) δ 7.68 (d, 1H, J ) 8.5 Hz), 7.64 (s, 1H), 7.17 (d, 1H, J ) 2.0 Hz), 6.94 (dd, 1H, J ) 8.5, 2.0 Hz), 6.90 (s, 1H), 5.70-5.50 (m, 2H), 1.67 (d, 3H, J ) 6.0 Hz). 13C NMR (DMSO-d6, 125 MHz) δ 159.6 (C), 157.8 (C), 157.0 (C), 155.9 (C), 153.1 (C), 128.7 (CH), 126.3 (CH), 126.2 (CH), 125.3 (C), 121.3 (C), 120.5 (CH), 118.6 (C), 114.7 (CH), 113.9 (CH), 102.4 (C), 98.7 (CH), 31.9 (CH2), 17.8 (CH3). HRMS (EI+) m/z 322.0847 ([M]+ C19H14O5, requires 322.0841). Methyl-3-[2,4-dimethoxy-5-(3-methyl-but-2-enyl)-phenyl]-2(2,4-dimethoxy-phenyl)-3-oxo-propionate (5). To a stirred solution of compound 11 (340 mg, 1.36 mmol) in CH2Cl2 (8 mL) under N2 (30) Coquerel, Y.; Rodriguez, J. Eur. J. Org. Chem. 2008, 112, 1125–1132. (31) For general experimental methods, see Supporting Information.

atmosphere, DMF (one drop) and oxalyl chloride (0.18 mL, 2.04 mmol) were added. The mixture was stirred for 3 h at rt, and then the solvent was removed under vacuum to yield acid chloride 7, which was directly used for the next reaction without further purification. A solution of compound 6 (430 mg, 2.1 mmol) in THF (5 mL) was added at -78 °C to a solution of LDA, prepared from n-BuLi (2.5 M in THF, 1.1 mL, 2.72 mmol) and i-Pr2NH (0.38 mL, 2.72 mmol) in THF (6 mL) at 0 °C under N2 atmosphere. This produced a yellowish colored anion that was stirred at -78 °C for 45 min, and then a solution of crude acid chloride 7 in THF (10 mL) was added dropwise. The reaction mixture was stirred at the same temperature for another 45 min and then at room temperature for 4 h. The reaction was quenched by the addition of 10% HCl (10 mL), THF was removed in vacuum, and the water layer was extracted with ethyl acetate (2 × 50 mL). The organic layer was washed with water (2 × 20 mL) and brine (20 mL), dried (Na2SO4), and concentrated. The crude product was purified by column chromatography to yield 5 (495 mg, 82%) as a yellowish oily compound. Rf ) 0.3 (50% ethyl acetate in hexane); νmax (CHCl3) cm-1 2359, 2341, 1738, 1666, 1604, 1507, 1463, 1270, 1211, 1029. 1 H NMR (CDCl3, 500 MHz) δ 7.72 (s, 1H), 7.01 (d, 1H, J ) 8.0 Hz), 6.41 (s, 1H), 6.40 (d, 1H, J ) 8.0 Hz), 6.30 (s, 1H), 5.91 (s, 1H), 5.22 (t, 1H, J ) 7.5 Hz), 3.82 (s, 3H), 3.76 (s, 3H), 3.75 (s, 3H), 3.72 (s, 3H), 3.70 (s, 3H), 3.18 (d, 2H, J ) 7.5 Hz), 1.68 (s, 3H), 1.64 (s, 3H). 13C NMR (CDCl3, 125 MHz) δ 193.0 (C), 171.1 (C), 162.6 (C), 160.3 (C), 159.6 (C), 157.9 (C), 132.7 (CH), 132.3 (CH), 130.3 (CH), 122.6 (C), 122.0 (C), 118.2 (C), 115.9 (C), 104.1 (CH), 98.5 (CH), 94.2 (CH), 57.3 (CH3), 55.6 (CH3), 55.5 (CH3), 55.4 (CH3), 55.3 (CH3), 52.2 (CH), 27.6 (CH2), 25.9 (CH3), 17.8 (CH3). HRMS (EI+) m/z 442.1993 ([M]+ C25H30O7, requires 442.1992). 1-Bromo-2,4-dimethoxy-5-(3-methyl-but-2-enyl)-benzene (10). Compound 9 (3.10 g, 10.47 mmol) was added to a solution of i-PrMgCl (1 M in THF, 13 mL, 13 mmol) at -10 °C under N2 atmosphere. After 45 min of stirring, prenyl bromide (1.3 mL, 11.5 mmol) was added dropwise, and the mixture was allowed to stir at the same temperature for 30 min and then at rt for 2 h. The reaction was quenched by the addition of 10% HCl (15 mL). THF was removed in vacuum, and the mixture was extracted with ethyl acetate (2 × 75 mL). The organic layer was washed with water (2 × 30 mL) and brine (20 mL), dried (Na2SO4), filtered, and concentrated. The resulting crude compound was purified by column chromatography to yield 10 (1.94 g, 65%) as a colorless liquid along with 550 mg of unreacted starting material 9. Rf ) 0.5 (5% ethyl acetate in hexane); νmax (CHCl3) cm-1 1599, 1500, 1463, 1295, 1208, 1029. 1H NMR (CDCl3, 500 MHz) δ 7.21 (s, 1H), 6.44 (s, 1H), 5.21 (t, 1H, J ) 7.0 Hz), 3.87 (s, 3H), 3.82 (s, 3H), 3.19 (d, 2H, J ) 7.0 Hz), 1.72 (s, 3H), 1.67 (s, 3H). 13C NMR (CDCl3, 125 MHz) δ 157.5 (C), 154.8 (C), 133.1 (C), 133.0 (CH), 124.0 (C), 122.2 (CH), 101.6 (C), 96.8 (CH), 56.6 (CH3), 55.9 (CH3), 27.6 (CH2), 26.0 (CH3), 17.9 (CH3). HRMS (EI+) m/z 284.0414 ([M]+ C13H17O2Br, requires 284.0412). 2,4-Dimethoxy-5-(3-methyl-but-2-enyl)-benzoic acid (11). nBuLi (2.5 M in THF, 2.1 mL, 5.34 mmol) was added dropwise to a stirred solution of 10 (1.27 g, 4.45 mmol) in THF (15 mL) at -78 °C under N2 atmosphere. After 30 min, CO2 gas was passed through the solution for 45 min, and then it was allowed to warm up to rt. THF was removed in vacuum, and the mixture was treated with saturated NaHCO3 solution (30 mL). The water layer was washed with ethyl acetate (2 × 20 mL) and then acidified with conc. HCl. The mixture was extracted with ethyl acetate (2 × 75 mL). The organic layer was washed with water (2 × 30 mL), brine (30 mL), dried (Na2SO4), filtered, and concentrated. Recrystallization from ethyl acetate produced 11 (930 mg, 84%) as a white solid. Rf ) 0.1 (50% ethyl acetate in hexane); mp: 98-99 °C; νmax (film): cm-1 1719, 1616, 1442, 1278,1168, 1020. 1H NMR (CDCl3, 500 MHz) δ 7.77 (s, 1H), 6.41 (s, 1H), 5.16 (t, 1H, J ) 7.0 Hz), 3.97 (s, 3H), 3.84 (s, 3H), 3.14 (d, 2H, J ) 7.0 Hz), 1.64 (s, 3H), J. Org. Chem. Vol. 74, No. 7, 2009 2753

Pahari and Rohr 1.60 (s, 3H). 13C NMR (CDCl3, 125 MHz) δ 166.2 (C), 162.6 (C), 158.5 (C), 133.6 (CH), 132.9 (C), 123.9 (CH), 121.6 (C), 108.9 (C), 94.5 (CH), 56.7 (CH3), 55.7 (CH2), 27.6 (CH3), 25.8 (CH3), 17.7 (CH3). HRMS (EI+) m/z 250.1206 ([M]+ C14H18O4, requires 250.1205). Methyl-3-(5-bromo-2,4-dimethoxy-phenyl)-2-(2,4-dimethoxyphenyl)-3-oxo-propionate (15). This compound was prepared as a yellowish oil (320 mg, 74%) from compound 13 and 6, following an analogous reaction sequence as described above for the synthesis of compound 5 (for the exact procedure, see Supporting Information). Rf ) 0.3 (50% ethyl acetate in hexane); mp 136-137 °C; νmax (film): cm-1 1730, 1668, 1613, 1590, 1508, 1465, 1334, 1271, 1213, 1155, 1021. 1H NMR (CDCl3, 500 MHz) δ 8.10 (s, 1H), 7.00 (d, 1H, J ) 9.0 Hz), 6.43 (s, 1H), 6.42 (d, 1H, J ) 9.0 Hz), 6.34 (s, 1H), 5.88 (s, 1H), 3.91 (s, 3H), 3.80 (s, 3H), 3.76 (s, 6H), 3.72 (s, 3H). 13C NMR (CDCl3, 125 MHz) δ 192.1 (C), 170.8 (C), 160.6 (C), 160.5 (C), 160.1 (C), 157.9 (C), 136.0 (CH), 130.4 (CH), 120.2 (C), 115.3 (C), 104.3 (CH), 103.2 (C), 98.7 (CH), 95.8 (CH), 57.3 (CH3), 56.6 (CH3), 55.7 (CH3), 55.6 (CH3), 55.4 (CH3), 52.4 (CH). HRMS (EI+) m/z 452.0477 ([M]+ C20H21O7Br, requires 452.0471). 2-Bromo-3,9-dihydroxy-benzo[4,5]furo[3,2-c]chromen-6-one (16). BBr3 (1 M in CH2Cl2, 1.72 mL, 1.72 mmol) was added at 0 °C to a solution of 15 (130 mg, 0.29 mmol) in CH2Cl2 (6 mL) under N2 atmosphere. The mixture was stirred at the same temperature for 10 h, and water (10 mL) was added. CH2Cl2 was removed in vacuum, and the mixture was heated to reflux for 1 h. The water layer was extracted with ethyl acetate (2 × 30 mL), and the organic layer was washed with water (2 × 15 mL) and brine (10 mL), dried (Na2SO4), and concentrated. Crude product was purified by column chromatography to get 16 (78 mg, 78%) as a white solid. Rf ) 0.3 (50% ethyl acetate in hexane); mp 362-364 °C; νmax (film) cm-1 1719, 1634, 1599, 1418, 1368, 1272, 1085. 1 H NMR (DMSO- d6, 500 MHz) δ 8.08 (s, 1H), 7.68 (d, 1H, J ) 8.5 Hz), 7.15 (d, 1H, J ) 2.0 Hz), 7.03 (s, 1H), 6.94 (dd, 1H, J ) 8.5, 2.0 Hz). 13C NMR (DMSO-d6, 125 MHz) δ 158.2 (C), 157.5 (C), 157.3 (2C, C), 156.1 (C), 153.4 (C), 125.0 (CH), 120.8 (CH), 114.5 (C), 114.2 (CH), 107.0 (C), 105.4 (C), 103.9 (CH), 102.8

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(C), 98.7 (CH). HRMS (EI+) m/z 345.9475 ([M]+ C15H7O5Br, requires 345.9477). 2-Bromo-3,9-bis-methoxymethoxy-benzo[4,5]furo[3,2-c]chromen-6-one (17). i-Pr2NEt (0.05 mL, 0.29 mmol) was added to a solution of 16 (40 mg, 0.11 mmol) in CH2Cl2 (6 mL) at rt. The mixture was stirred for 15 min, and then MOMCl (0.02 mL, 0.25 mmol) was added. The reaction was stirred for 10 h and quenched by the addition of water (5 mL). The aqueous layer was extracted with ethyl acetate (2 × 20 mL) and the organic layer was washed with water (2 × 10 mL) and brine (10 mL), dried (Na2SO4), and concentrated. The crude product was purified by column chromatography to get 17 (35 mg, 65%) as a white solid. Rf ) 0.8 (30% ethyl acetate in hexane); mp 212-214 °C; νmax (film) cm-1 1758, 1629, 1490, 1353, 1257, 1163, 1072, 957. 1H NMR (CDCl3, 500 MHz) δ 8.14 (s, 1H), 7.94 (d, 1H, J ) 8.5 Hz), 7.35 (s, 1H), 7.29 (s, 1H), 7.14 (d, 1H, J ) 8.5 Hz), 5.33 (s, 2H), 5.25 (s, 2H), 3.53 (s, 3H), 3.51 (s, 3H). 13C NMR (CDCl3, 125 MHz) δ 159.0 (C), 158.0 (C), 157.2 (C), 156.5 (C), 156.3 (C), 153.9 (C), 125.6 (CH), 122.0 (CH), 117.6 (C), 115.5 (CH), 109.3 (C), 108.2 (C), 104.7 (CH), 102.7 (C), 99.9 (CH), 95.5 (CH2), 95.2 (CH2), 56.9 (CH3), 56.4 (CH3). HRMS (EI+) m/z 433.9992 ([M]+ C19H15O7Br, requires 434.0001).

Acknowledgment. This work was supported by a grant from the National Institutes of Health (AT002890) to J.R. Authors are thankful to Professor Gregory I. Elliott for the use of the microwave instrument and also for valuable advice, the NMR core facility and mass spectrometry facility of the University of Kentucky, as well as the Department of Chemistry at the University of Kentucky for letting us use their instruments. Supporting Information Available: General experimental methods, experimental details, and characterization data for compounds 9, 12, 13, 15, 18, 19, 21-23, and copies of 1H and 13 C NMR spectra of all new compounds. This material is available free of charge via the Internet at http://pubs.acs.org. JO8025884